アポトーシス機構の解析とその発がん、制がん研究への応用

• Project/Area Number: 08265107
• Research Category: Grant-in-Aid for Scientific Research on Priority Areas (A)
• Allocation Type: Single-year Grants
• Research Institution: National Cancer Center Research Institute and Research Center for Innovative Oncology, National Cancer Center Hospital East
• Principal Investigator: 口野 嘉幸 国立がんセンター研究所, 生物物理部, 部長 (60124418)
• Co-Investigator(Kenkyū-buntansha): 中島 琢磨 東京理科大学, 工学部, 助手 (90256678) ; 田沼 靖一 東京理科大学, 薬学部, 教授 (10142449) ; 三浦 正幸 大阪大学, 医学部, 助教授 (50202338) ; 藤田 直也 東京大学, 分生研, 助手 (20280951) ; 松沢 昭雄 東京大学, 医科研, 助教授 (50012745)
• Project Period (FY): 1996 – 1998
• Project Status: Completed (Fiscal Year 1998)
• Budget Amount: ¥84,000,000 (Direct Cost: ¥84,000,000); Fiscal Year 1998: ¥28,000,000 (Direct Cost: ¥28,000,000); Fiscal Year 1997: ¥28,000,000 (Direct Cost: ¥28,000,000); Fiscal Year 1996: ¥28,000,000 (Direct Cost: ¥28,000,000)
• Keywords: Myc / アデノウイルスE1A / 制がん剤 / Bax / FasL / アポトーシスシグナル / カスパーゼ3様プロテアーゼ / アポトーシス耐性 / アポトーシス / LPS / Pim-1キナーゼ / リンパ腫 / ICE / CED-3 family proteases / myc関連遺伝子 / Fas / エンドトキシンLPS / トポイソメラーゼIIα / アポトーシスエンドヌクレアーゼ

Research Abstract

アポトーシスはがん遺伝子、がん抑制遺伝子をはじめ、抗がん剤や放射線など細胞の内外に存在する様々な因子によって誘導・制御されている。本研究班ではこれら性質の異なった刺激によって活性化されるアポトーシスシグナルの伝達機構を各班員が独自に開発した実験系を用いて解析した。その結果c-mycやアデノウイルスE1Aなどのがん遺伝子をはじめ、がん抑制遺伝子としてのs-mycやp53の標的遺伝子であるbaxの発現、さらには制がん剤処理など、様々な刺激によって誘導されるアポトーシスにおいてもカスパーゼ3様プロテアーゼの活性化が共通して要求されることを見いだし、カスパーゼ3様プロテアーゼがアポトーシス実行において中心的役割を果たしていることを明らかにした。またがん遺伝子やがん抑制遺伝子の変異や発現亢進、がん遺伝子との協調発現によって細胞のがん化に関与する遺伝子の発現がしばしば細胞にアポトーシス耐性を与えていることを明らかにした。これら一連の研究成果は抗がん剤等のアポトーシスにおける作用点や作用機構を知る手掛かりとなり、アポトーシスを標的とした新たながんの治療法の開発に利用できるものと期待される。一方s-MycやFasLを発現させたがん細胞の移植実験からアポトーシスを介したlocal inflammationが宿主の抗腫瘍免疫能の活性化を惹き起こすことを見出し、アポトーシスを利用したがんに対する免疫療法の可能性を示唆した。最後に本研究において活性型Rasがアポトーシスとは異なったタイプの細胞死を誘導することを明らかにした。この細胞死はカスパーゼの活性化を要求せず、p53やBcl-2による制御も受けない。このような特徴をもつカスパーゼ非依存的細胞死誘導システムは、多くのアポトーシス耐性を獲得したがんの治療法として非常に有用であると思われる。この発見はまたこれまで不明な点が多かったプログラム細胞死の全体像を分子レベルで理解していく上で非常に意義があると思われる。

Research Products

• [Publications] K.Hanaki,et al.: "Two different reactions involved in the primer/template-independent polymerization of dA TP and dTTP by Taq DNA polymerase." Biochem.Biophys.Res.Commun.244. 210-219 (1998)
• [Publications] K.Mishima,et al.: "Heparin-binding epidermal growth factor-like growth factor stimulates mitogenic signaling and is highly expressed in human malignant gliomas." Acta Neuropathologica. 96. 322-328 (1998)
• [Publications] A.Sugiyama,et al.: "Molecular cloning and chromosomal mapping of mouse intronless myc gene acting as a potent apoptosis inducer." Gene. 226. 273-283 (1999)
• [Publications] S.Chi,et al.: "Oncogenic Ras triggers cell suicide through the activation of a caspase-independent cell death program in human cancer cell." Oncogene,in press.
• [Publications] C.Kitanaka,et al.: "Caspase-independent cell death with necrotic morphology: New insight into the signaling pathway for programmed cell death." Cell Death & Differ.,in press.
• [Publications] T.Araki,et al.: "Caspase activity is required for nephrogenesis in the developing mouse metanephros." Exp.Cell Res.,in press.
• [Publications] A.Suzuki,et al.: "Caspase-3 inactivation to suppress Fas-mediated apoptosis: identification of binding domain with p21 and ILP and inactivation machinery by p21." Oncogene,in press.
• [Publications] H.Kanuka,et al.: "Pro-apoptotic activity of C.elegans CED-4 in Drosophila: Implicated mechanisms for caspase activation." Proc.Natl.Acad.Sci.USA. 96. 145-150 (1999)
• [Publications] A.Suzuki,et al.: "Resistance to Fas-Mediated apoptosis: activation of caspase-3 is regulated by cell cycle regulator p21 WAF and IAP gene family ILP." Oncogene. 17. 931-939 (1998)
• [Publications] S.Wang,et al.: "Murine caspase-11,an ICE interacting protease,is essential for activation of ICE." Cell. 92. 501-509 (1998)
• [Publications] N.Fujita,et al.: "Acceleration of apoptotic cell death after the cleavage of Bcl-XL protein by caspase-3-like proteases." Oncogene. 17. 1295-1304 (1998)
• [Publications] N.Fujita,et al.: "Involvement of Bcl-2 cleavage in the acceleration of VP-16-induced U937 cell apoptosis." Biochem.Biophys.Res.Commun.246. 484-488 (1998)
• [Publications] Y.Morinaga,et al.: "Suppression of interleukin-11-mediated bone resorption by cyclooxygenases inhibitors." J.Cell.Physiol.175. 247-254 (1998)
• [Publications] Y.Zhang,et al.: "Caspase-mediated cleavage of p21 Waf 1/Cip 1 converts cancer cells from growth arrest to undergoing apoptosis." Oncogene,in press.
• [Publications] K.Kage,et al.: "Basic fibroblast growth factor induces cyclooxygenase-2 expression in endothelial cells derived from bone." Biochem.Biophys.Res.Commun.,in press.
• [Publications] D.Shiokawa,et al.: "Molecular cloning and expression of a cDNA encoding an apoptotic endonuclease DNaseγ." Biochem.J.332. 713-720 (1998)
• [Publications] D.Shiokawa,et al.: "cDNA cloning of human DNaseγ: chromosomal localization of its gene and enzymatic properties of recombinant protein." Apoptosis. 3. 89-95 (1998)
• [Publications] K.Nishimura,et al.: "Presence of DNaseγ-like endonuclease in nuclei of neuronal differentiated pC12 cells." Apoptosis. 3. 97-103 (1998)
• [Publications] F.Uchiumi,et al.: "Caracterization of telomere-binding activity of replication factor C large subunit p140." Nucleic acids Res.,in press.
• [Publications] N.Matsumura,et al.: "Purification and properties of an NAD+ glycohydrolase from bovine brain." Eur.J.Biochem,in pres.
• [Publications] I.Iwakura,et al.: "The development of autoimmune in flammatory arthropathy in mice transgenic for the HTVL-1 env-pX region is not dependent on H-2 haplotypes but dependent on expression of Fas antigen." J.Immunol.161. 6592-6598 (1998)
• [Publications] H.Nagase,et al.: "Novel mutant mice secreting soluble CD4 without expression of membrane-bound CD4" Eur.J.Immunol.28. 403-412 (1998)
• [Publications] K.Seino,et al.: "Chemotactic activity of soluble Fas ligand against polymorphonuclear leukocytes." J.Immunol.161. 4484-4488 (1998)
• [Publications] M.Shimizu,et al.: "Amtitumor activity exhibited by Fas ligand (CD95L)overexpressing on lymphoid cells against Fas+ tumor cells." Cancer Imunol.Immunother.47. 143-148 (1998)
• [Publications] M.Shimizu,et al.: "Induction of antitumor immunity with Fas/APO-1 ligand(CD95L)-transfected neuroblastoma Neuro-2α cells." J.Immunol.,in press.
• [Publications] N.Ohi,et al.: "A novel adenovirus E1B19K-binding protein B5 inhibits apoptosis induced by Nip3 by forming a heterodimer through the C-terminal hydrophobic region." Cell Death & Differ.,in press.
• [Publications] T.Nakajuma,et al.: "Suppression of adenovirus E1A-induced apoptosis by mutated p53 is overcome by coexpression with Id proteins." Proc.Nat1.Acad.Sci.USA. 95. 10590-10595 (1998)
• [Publications] T.Nakajima,et al.: "Stabilization of p53 adenovirus E1A occurs through its amino-terminal region by modification of the ubiquitin-proteasome pathway." J.Biol.Chem.273. 20036-20045 (1998)
• [Publications] M.Suzuki,et al.: "Induction of Sp1 in defferentiating human embryonal carcinoma cells triggers transcription of the fibronectin gene." Mol.Cell.Biol.18. 3010-3020 (1998)
• [Publications] M.Shimizu,et al.: "Activation of the rat cyclin A promoter by ATF2 and Jun family members and its suppression by ATF4." Exp.Cell Res.239. 93-103 (1998)
• [Publications] S.Kagaya, et al.: "A functional role for death proteases in s-Myc-and c-Myc-mediated apoptosis." Mol.Cell.Biol.17. 6736-6745 (1997)
• [Publications] T.Mochizuki, et al.: "Pim-1 kinase stimulates c-Myc-mediated death signaling upstream of caspase-3(CPP-32)-like protease activation." Oncogene. 15. 1471-1480 (1997)
• [Publications] C.Kitanaka, et al.: "Caspase-dependent apoptosis of COS-7 cells induced by Bax overexpression:differential effects of Bcl-2 and Bcl-X_T on Bax-induced caspase activation and apoptosis." Oncogene. 15. 1763-1772 (1997)
• [Publications] H.Hisahara, et al.: "ICE/CED-3 family execute oligodendrocytes apoptosis by tumor necrosis factor." J.Neurochem.69. 10-20 (1997)
• [Publications] N.Fujita, et al.: "Aggregation of Thy-1 glycoprotein induces thymocyte apoptosis through activation of CPP32-like proteases." Exp.Cell Res.232. 400-406 (1997)
• [Publications] Y.Kato, et al.: "Suppression of experimental lung colonization of mouse colon adenocarcinoma 26 in vivo by an anti-idiotype monoclonal antibody recognizing a platelet surface molecule." Cancer Res.57. 3040-3045 (1997)
• [Publications] T.Nakajima, et al.: "Induction of ubiquitin conjugation enzyme activity for degradation of topoisomerase IIα during adenovirus E1A induced apoptosis." Biochem.Biophys.Res.Commun.239. 823-829 (1997)
• [Publications] H.Nagase, et al.: "Novel mutant mice secreting soluble CD4 without expression of membrane-bound CD4." Eur.J.Immunol.,. (in press). (1997)
• [Publications] H.Nakano, et al.: "Genetic defect in T lymphocyte homing into peripheral lymph nodes." Eur.J.Immunol.27. 215-221 (1997)
• [Publications] D.Shiokawa, et al.: "Inhibitors of poly(ADP-ribose)polymerase suppress nuclear fragmentation and apoptotic-body formation during apoptosis in HL-60 cells." FEBS Lett.413. 99-103 (1997)
• [Publications] A.Nishikawa, et al.: "Occurence of DNaseγ-like apoptotic endonucleases in hematopoietic cells in X enopus Laevis and their relation to metamorphosis." Biochem.Biophys.Res.Commun.231. 305-308 (1997)
• [Publications] Y.Kuchino,et al.: "Myc-Mediated Apoptosis." Prog.Mol.Subcell.Biol.16. 104-129 (1996)
• [Publications] A.Asai,et al.: "Application of the Apoptotic gene to gene therapy of malignant gliomas." Brain Tumor Research and Therapy. 397-407 (1996)
• [Publications] Y.Kuchino,et al.: "Apoptosis in Cancer." HUMAN CELL. 9. 223-228 (1996)
• [Publications] S.Wang,et al.: "Identification and characterization of Ich-3,a member of the ICE/Ced-3 family and an upstream regulator of ICE." J.Biol.Chem.271. 20580-20587 (1996)
• [Publications] Y-K.Jung,et al.: "Suppression of interleukin-1 β-converting enzyme-mediated cell death by insulin-like growth factor." J.Biol.Chem.271. 5112-5117 (1996)
• [Publications] K.Noguchi,et al.: "Chromosome 22 complements apoptosis in Fas-and TNF-resistant mutant UK110 cells." Oncogene. 13. 39-46 (1996)
• [Publications] S-H.Lee,et al.: "Interleukin-7 inhibits apoptosis of T-lymphoma cells by both inducing the bcl-2 expression and suppressing the ICE-family proteases activation." Oncogene. 13. 2131-2139 (1996)
• [Publications] T.Nakajima,et al.: "Degradation of topoisomerase II α during adenovirus E1A-induced apoptosis is mediated by the activation of the ubiquitin proteolysis system." J.Biol.Chem.271. 24842-24849 (1996)
• [Publications] K.Tanuma,et al.: "Molecular mechanisms of apoptosis." Apoptosis in Normal Development and Cancer. 39-59 (1996)
• [Publications] D.Shiokawa,et al.: "DNA fragmentation during thymic apoptosis is catalyzed by DNase γ." Apoptosis. 1. 147-152 (1996)
• [Publications] A.Suzuki,et al.: "Involvement of Fas in regression of vaginal epithelia after ovariectomy and during an estrous cycle." EMBO. 15. 211-215 (1996)
• [Publications] A.Suzuki,et al.: "Down regulation of cl-2 is the first step of Fas-mediated apoptosis of male reproductive tract." Oncogene. 13. 31-37 (1996)